The present invention relates to analogs of tetrapeptides that inhibit protein:prenyl transferase. Protein:prenyl transferases are understood to comprise protein:farnesyl transferases, protein:geranylgeranyl transferases and any other enzymes capable of transferring prenyl groups to proteinaceous substrates.
The post-translational modification of mammalian cell proteins involve thioether derivatisation of carboxyl-terminal cysteine residues by mevalonate-derived isoprenyl groups. Several of these prenylated proteins were identified as belonging to groups of related proteins: e.g. the nuclear lamins, low molecular weight GTP binding proteins, such as the ras-oncogene proteins and heteromeric G proteins (Schxc3xa4fer et al., Science 245 (1989) 379-385). The prenyl group that was attached to a protein was identified as either farnesyl (C15) or geranylgeranyl (C20), probably depending on the recognition of the C-terminal amino acid sequence of the proteins involved, Lamins and p21ras proteins, 188 or 189 amino acid proteins which possess the consensus CAAX motif (C=cysteine, A=any amino acid having an aliphatic side chain and X=methionine, serine, glutamine or alanine) at the C terminus, are farnesylated, while several members of the rab proteins having C-terminal CC/CXC motifs, and of the heteromeric G-protein xcex3-subunits are geranylgeranylated.
The prenylation of these proteins seems to play a role in their association with membranes and nuclear envelopes, where they are processed further and/or perform their function. This was shown for example by blocking the mevalonate synthesis by HMG-CoA reductase inhibitors, which prevented proteolytic processing of the lamin A precursor (Beck et al., J. Cell. Biol. 110 (1990) 1489-1499) or resulted, in other studies, in the accumulation of non-prenylated p21ras precursor and the loss of transforming activity of oncogenic ras proteins. A review of the post-translational modification of proteins by isoprenoids in mammalian cells is given by Maltese W.A. in FASEB J. 4 3319-3329 (1990). The latter observation triggered the search for specific inhibitors of the farnesylation of p21ras in order to prevent its action in cells, where overexpression of this protein leads to tumour development, such as in colon carcinomas.
G proteins play a role in the receptor-mediated transduction of signals (such as growth modulation signals) over the plasma membrane, and other prenylated proteins, not yet identified, may have a function in cell cycle progression. There is some evidence as well that GTP binding proteins are involved in the regulation of intracellular protein traffic and secretion. There is even some suggestion that prenylated proteins play a role in the translational control of HMG-CoA reductase, the rate limiting enzyme of the isoprene and subsequent cholesterol synthesis.
The enzymes involved in the protein prenylation process, protein:prenyl transferases, are reported to use all-trans-farnesyl pyrophosphate (FPP) as a substrate for the addition of the farnesyl group to the protein. FPP is a substrate in the productoin of geranylgeranyl pyrophosphate, which is subsequently used in the synthesis of geranylgeranylated proteins. EP-A-540782 describes inhibitors for protein:farnesyl transferase based on prenyl pyrophosphate analogues.
A review of new therapeutic methods based on Ras farnesyltransferase and inhibitors thereof are described in Leonard in J. Med. Chem. 40 (1997), 2971-2990. A first group of potential inhibitors is based on the CAAX tetrapeptide motive of the ras proteins. The proposed analogues include CVFM (SEQ ID NO:1) as well as structures having reduced amide bonds and further deviations from pure tetrapeptide structures. Nigam et al (J. Biol. Chem. 268 (1993), 20695-98) reported that Cys-NH-CH2-mC6H4-CO-Met inhibits Ras farnesyl transferase from human colon carcinoma with an IC50 of 60 nM.
Novel tetrapeptide analogs have been found according to the invention, which are effective inhibitors of protein:prenyl transferases such as protein:farnesyl transferase and protein:geranylgeranyl transferases. The analogs are defined in the appending claims with reference to formula 1. These inhibitors are preferably not substrates for protein:prenyl transferases. The novel tetrapeptide s contain conformationally restricted dipeptide isoster moieties which have the potential to be unsusceptible towards enzymatic degradation.